Recombinant DNA technology has made it possible to express genes of one organism within another. The prior art shows that several virus groups including the papovaviruses, papilloma viruses, adenoviruses, and retroviruses have been employed as eukaryotic molecular cloning and expression vectors. The relatively small sizes of these virus genomes have facilitated the in vitro construction of recombinant DNA molecules. However, they generally exhibit a limited host range, provide severe limitations on the amounts of DNA that can be accommodated and suffer loss of infectivity upon insertion of foreign DNA. Although genetic engineering of larger viruses, such as poxviruses, is more difficult, such vectors could have the advantage of greater capacity and potential of retaining infectivity in a wide range of host cells. For poxviruses such as vaccinia virus, such recombinants may lead to the development of live virus vaccines.
Since vaccinia virus is the best studied member of the poxvirus group, it will be described here. Vaccinia virus has a very broad host range in vitro and in vivo and has been used world-wide as an effective vaccine against variola, a related poxvirus that causes smallpox. Vaccinia is a large virus containing a linear double-stranded DNA genome with a molecular weight of about 122 million, equivalent to more than 180,000 base pairs. The virus uses its own enzymes for transcription and replication within the cytoplasm of infected cells. Nucleotide sequence data indicate that the transcriptional regulatory signals encoded in the vaccinia virus genome are distinct from those used by eukaryotic cells. The invention described here takes into account both the large size of the poxvirus genome and its unique transcriptional regulatory signals.
References which relate to the subject invention are Venkatesan, Baroudy and Moss, Cell 125: 805–813 (1981); Venkatesan, Gershowitz and Moss, J. Virol. 44: 637–646 (1982); Bajszar, Wittek, Weir and Moss, J. Virol. 45: 62–72 (1983); Weir and Moss, J. Virol. 46: 530 (1983); Moss, Winters and Cooper, J. Virol. 40: 387–395 (1981); Panicali and Paoletti, Proc. Natl. Acad. Sci. USA 79: 4927–4931 (1982); Mackett, Smith and Moss, Proc. Natl. Acad. Sci. USA 79: 7415 (1982); Cohen and Boyer, U.S. Pat. No. 4,237,224; Valuenzuela et al., Nature 298: 347–350 (1982); Moriarity et al., Proc. Natl. Acad. Sci. USA 78: 2606–2610 (1981); Liu et al., DNA 1: 213–221 (1982); Weir, Bajszar and Moss, Proc. Natl. Acad Sci USA 79: 1210 (1982).